The general objective of the proposed research is to use the Escherichia coli and Salmonella typhimurium omega proteins as model systems to investigate enzymatic control of DNA topology. The specific aim is to understand the molecular basis and physiological role(s) of DNA topoisomerases as typified by these proteins. The omega protein (DNA topoisomerase) of S. typhimurium will be purified to homogeneity for detailed physical and chemical analysis. The S. typhimurium and E. coli enzymes will be compared with respect to physical and chemical properties, and common features will be related to catalytic function. Binding of homogeneous omega proteins to DNA molecules will be investigated, to obtain thermodynamic and kinetic parameters characterizing the interactions of these proteins with DNA. Procedures will be derived to separate and distinguish the two enzymes from mixtures of E. coli and S. typhimurium cell extracts. The structural gene of the E. coli omega protein will be mapped by means of interspecies genetic hybrid strains. S. Typhimurium strains carrying small regions of the E. coli chromosomme on E. coli F' plasmids will be examined for the presence of the E. coli omega protein. Once the E. coli omega gene has been located and other closely linked genes identified, localized mutagenesis will be employed to generate mutations within this region. Mutants with altered omega activity will be isolated, and used to study the physiological role(s) of the E. coli omega protein.